Toastier nest temperatures, rather than sex chromosomes, turn baby turtles female. Now, a genetic explanation for how temperature determines turtles’ sex is emerging: Scientists have identified a temperature-responsive gene that sets turtle embryos on a path to being either male or female. When researchers dialed down that gene early in development, turtle embryos incubating at the cooler climes that would normally yield males turned out female instead, researchers report in the May 11 Science.

Scientists have struggled since the 1960s to explain how a temperature cue can flip the sex switch for turtles and other reptiles (SN Online: 1/8/18). That’s partly because gene-manipulating techniques that are well-established in mice don’t work in reptiles, says study coauthor Blanche Capel, a developmental biologist at Duke University School of Medicine. Previous studies showed certain genes, including one called Kdm6b, behaving differently in developing male and female turtles. But until recently, nobody had been able to tweak those genes to directly test which ones controlled sex.

“This is the first venture down that path,” says Clare Holleley, an evolutionary geneticist at the Australian National Wildlife Collection in Canberra who wasn’t part of the study. “It's really quite a breakthrough.”

In the new study, Capel’s lab collaborated with a group of Chinese researchers led by Chutian Ge of Zhejiang Wanli University in Ningbo. Ge’s team recently developed a way to lessen the activity of particular reptilian genes by injecting viruses bearing snippets of artificial RNA into developing eggs.

The researchers used the technique to weaken the effects of the Kdm6b gene in the embryos of red-eared slider turtles (Trachemys scripta elegans) before the gonads formed, then tracked the embryos’ development at 26° Celsius.

“To my delight, it resulted in complete sex reversal,” Capel says. That temperature should have yielded all male turtles. Instead, in two separate experiments done with different gene-silencing viruses, 80 and 87 percent of the surviving embryos became female.

Still, something as complex as sex determination can’t be boiled down to a single gene. Kdm6b controls a gene called Dmrt1, which had already been shown to direct male development, Capel’s team also found. And while Kdm6b does behave differently as temperatures rise, it doesn’t show the same response in all tissues. That suggests that the gene doesn’t directly sense temperature, but is instead receiving messages from some higher-up gene that reacts directly to temperature and directs Kdm6b’s behavior in different tissues, the researchers propose.

Whether Kdm6b plays the same role in other reptiles remains to be seen. A 2017 study in Science Advances coauthored by Holleley found that the gene influenced bearded dragons’ sexual fate (SN Online: 6/14/17). But other genes in the same family, Jumonji genes, are also known to influence development in both reptiles and mammals. And those genes might not work exactly the same way in other reptiles.

“There's this huge diversity of sex determining modes in reptiles,” Holleley says. Even if Kdm6b is an important switch in other reptiles, “the genes that Jumonji genes are activating are probably going to be different in every reptile.”

There are other wrinkles, too. “This is really exciting finding, but we need to remember that everything in a lab is controlled,” says Itzel Sifuentes-Romero of Florida Atlantic University in Boca Raton. Wild turtle eggs are subject to fluctuations in temperature and moisture as they incubate, she says, which means the signals that temperature-sensitive genes are receiving are far more muddled.